Horizontal gene transfer in microbial genome evolution

Horizontal gene transfer is the collective name for processes that permit the exchange of DNA among organisms of different species. Only recently has it been recognized as a significant contribution to inter-organismal gene exchange. Traditionally, it was thought that microorganisms evolved clonally, passing genes from mother to daughter cells with little or no exchange of DNA among diverse species. Studies of microbial genomes, however, have shown that genomes contain genes that are closely related to a number of different prokaryotes, sometimes to phylogenetically very distantly related ones. (Doolittle et al., 1990, J. Mol. Evol. 31, 383-388; Karlin et al., 1997, J. Bacteriol. 179, 3899-3913; Karlin et al., 1998, Annu. Rev. Genet. 32, 185-225; Lawrence and Ochman, 1998, Proc. Natl. Acad. Sci. USA 95, 9413-9417; Rivera et al., 1998, Proc. Natl. Acad. Sci. USA 95, 6239-6244; Campbell, 2000, Theor. Popul. Biol. 57 71-77; Doolittle, 2000, Sci. Am. 282, 90-95; Ochman and Jones, 2000, Embo. J. 19, 6637-6643; Boucher et al. 2001, Curr. Opin., Microbiol. 4, 285-289; Wang et al., 2001, Mol. Biol. Evol. 18, 792-800). Whereas prokaryotic and eukaryotic evolution was once reconstructed from a single 16S ribosomal RNA (rRNA) gene, the analysis of complete genomes is beginning to yield a different picture of microbial evolution, one that is wrought with the lateral movement of genes across vast phylogenetic distances. (Lane et al., 1988, Methods Enzymol. 167, 138-144; Lake and Rivera, 1996, Proc. Natl. Acad. Sci. USA 91, 2880-2881; Lake et al., 1999, Science 283, 2027-2028).     

Protein sequential resonance assignments by combinatorial enumeration using 13C alpha chemical shifts and their (i,i-1) sequential connectivities

The need for the structural characterization of proteins on a genomic scale has brought with it demands for new technology to speed the structure determination process. In NMR, one bottleneck is the sequential assignment of backbone resonances. In this paper, we explore the computational complexity of the sequential assignment problem using only ¹³C chemical shift data and C (i,i–1) sequential connectivity information, all of which can potentially be obtained from a single three-dimensional NMR spectrum. Although it is generally believed that there is too much ambiguity in such data to provide sufficient information for sequential assignment, we show that a straightforward combinatorial search algorithm can be used to find correct and unambiguous sequential assignments in a reasonable amount of CPU time for small proteins (approximately 80 residues or smaller) when there is little missing data. The deleterious effect of missing or spurious peaks and the dependence on match tolerances is also explored. This simple algorithm could be used as part of a semi-automated, interactive assignment procedure, e.g., to test partial manually determined solutions fo uniqueness and to extend these solutions.     

Short inverse complementary amino acid sequences generate protein complexity

Inversions of short genomic sequences play a central role in the generation of protein complexity. More than half of the 1300 motifs registered in ProSite have protein inverse complementary sequences (princoms) among proteins registered in SwissProt. The observed number of princoms occurrences exceeds by far the expected number (p < 10(-10)). Princoms often endow their host proteins with a whole new range of biochemical and physiological capabilities, including the possibility of intramolecular and intermolecular disulfide bond formation. These results support the idea that, like the duplications, the inversions of small genomic fragments have been a fundamental mechanism for shaping genomes.     

Cre-mediated recombination in the skin melanocyte lineage

Organ-specific expression of a Cre recombinase allows the analysis of gene function in a particular tissue or cell type. Using a 6.1 kb promoter from the mouse tyrosinase gene, we generated and characterized two lines of transgenic mice that express Cre recombinase in melanoblasts. Utilizing a Cre-responsive reporter mouse strain, genetic recombination was detected in the melanoblasts of the skin from embryonic day 11.5. In addition, Cre-expression was detected in the skin and eyes of mice. Cre transgene activity was occasionally detected in the brain and peripheral nerves but not in other tissues. When Tyr::Cre mice were crossed with mice carrying a homozygous loxP conditional mutation for the insulin-like growth factor receptor gene (Igf1r), Cre-melanoblast-specific recombination pattern was confirmed and no abnormal phenotype was observed. In conclusion, Tyr::Cre transgenic mice provide a valuable tool to follow the cell lineage and to examine gene function in melanocyte development and transformation.     

Conditional knockout of the mouse NADPH-cytochrome p450 reductase gene

NADPH-cytochrome P450 reductase (CPR or POR) is the obligatory electron donor for all microsomal cytochrome P450 (CYP or P450)-catalyzed monooxygenase reactions. Disruption of the mouse Cpr gene has been reported to cause prenatal developmental defects and embryonic lethality. In this study, we generated a mouse model with a floxed Cpr allele (termed Cpr(lox)). Homozygous Cpr(lox) mice are fertile and without any histological abnormality or any change in CPR expression. The floxed Cpr allele was subsequently deleted efficiently by crossing Cpr(lox) mice with transgenic mice having liver-specific Cre expression (Alb-Cre); the result was a decrease in the level of CPR protein in liver microsomes. The Cpr(lox) strain will be valuable for conditional Cpr gene deletion and subsequent determination of the impact of CPR loss on the metabolism of endogenous and xenobiotic compounds, as well as on postnatal development and other biological functions.     

The conservation of redundancy in genetic systems: effects of sexual and asexual reproduction

The relationship between probability of survival and the number of deleterious mutations in the genome is investigated using three different models of highly redundant systems that interact with a threatening environment. Model one is a system that counters a potentially lethal infection; it has multiple identical components that act in sequence and in parallel. Model two has many different overlapping components that provide three-fold coverage of a large number of vital functions. The third model is based on statistical decision theory: an ideal detector, following an optimum decision strategy, makes crucial decisions in an uncertain world. The probability of a fatal error is reduced by a redundant sampling system, but the chance of error rises as the system is impaired by deleterious mutations. In all three cases the survival profile shows a synergistic pattern in that the probability of survival falls slowly and then more rapidly. This is different than the multiplicative or independent survival profile that is often used in mathematical models. It is suggested that a synergistic profile is a property of redundant systems. Model one is then used to study the conservation of redundancy during sexual and asexual reproduction. A unicellular haploid organism reproducing asexually retains redundancy when the mutation rate is very low (0001 per cell division), but tends to lose high levels of redundancy if the mutation rate is increased (001 to 01 per cell division). If a similar unicellular haploid organism has a sexual phase then redundancy is retained for mutation rates between 0001 and 01 per cell division. The sexual organism outgrows the asexual organism when the above mutation rates apply. If they compete for finite resources the asexual organism will be extinguished. Variants of the sexual organism with increased redundancy will outgrow those with lower levels of redundancy and the sexual process facilitates the evolution of more complex forms. There is a limit to the extent that complexity can be increased by increasing the size of the genome and in asexual organisms this leads to progressive accumulation of mutations with loss of redundancy and eventual extinction. If complexity is increased by using genes in new combinations, the asexual form can reach a stable equilibrium, although it is associated with some loss of redundancy. The sexual form, by comparison, can survive, with retention of redundancy, even if the mutation rate is above one per generation. The conservation and evolution of redundancy, which is essential for complexity, depends on the sexual process of reproduction.     

Conditional ablation of beta1 integrin in skin. Severe defects in epidermal proliferation, basement membrane formation, and hair follicle invagination

The major epidermal integrins are alpha3beta1 and hemidesmosome-specific alpha6beta4; both share laminin 5 as ligand. Keratinocyte culture studies implicate both integrins in adhesion, proliferation, and stem cell maintenance and suggest unique roles for alphabeta1 integrins in migration and terminal differentiation. In mice, however, whereas ablation of alpha6 or beta4 results in loss of hemidesmosomes, epidermal polarity, and basement membrane (BM) attachment, ablation of alpha3 only generates microblistering due to localized internal shearing of BM. Using conditional knockout technology to ablate beta1 in skin epithelium, we have uncovered biological roles for alphabeta1 integrins not predicted from either the alpha3 knockout or from in vitro studies. In contrast to alpha3 null mice, beta1 mutant mice exhibit severe skin blistering and hair defects, accompanied by massive failure of BM assembly/organization, hemidesmosome instability, and a failure of hair follicle keratinocytes to remodel BM and invaginate into the dermis. Although epidermal proliferation is impaired, a spatial and temporal program of terminal differentiation is executed. These results indicate that beta1's minor partners in skin are important, and together, alphabeta1 integrins are required not only for extracellular matrix assembly but also for BM formation. This, in turn, is required for hemidesmosome stability, epidermal proliferation, and hair follicle morphogenesis. However, beta1 downregulation does not provide the trigger to terminally differentiate.     

Male mating strategies and the mating system of great-tailed grackles

Great-tailed grackles (Quiscalus mexicanus) are sexually dimorphic,dichromatic, colonially nesting blackbirds. In this study, males pursued three basic types of conditional mating strategies,each of which employed a different set of mating tactics. Territorialmales defended one or more trees in which several females nested.They achieved reproductive success by siring the offspringof their social mates and through extrapair fertilization.Resident males lived in the colony but did not defend territoriesor have social mates. Transient males passed through the colony, staying no more than a few days, and probably visited more thanone colony. Residents appeared to queue for access to territories,but transients did not. Residents and transients gained allpaternity through extrapair fertilizations and provided noparental care. Territorial males sired the majority of offspring,but residents and transients also sired small numbers of nestlings. Territorial males were larger and had longer tails than nonterritorialmales. The number of social mates was related to body size,and males that sired nestlings were heavier and had longertails than males with no genetic reproductive success. Malesthat gained paternity through extrapair fertilization wereheavier and had longer tails than males that did not. The matingsystem of great-tailed grackles can best be categorized as "non-faithful-female frank polygyny."     

The Industrial Ecology of Emerging Technologies

The modern world increasingly reflects human activities, to the point that many scientists are referring to this era as the "Anthropocene," the Age of Humans. A major domain of human activity involves sociotechnical systems, which can be characterized as occurring in constellations of coevolving technological, cultural, institutional, economic, and psychological systems lasting over many decades. The current constellation, still in its early stages of development, brings together five powerful technology systems—nanotechnology, biotechnology, robotics, information and communication technology, and cognitive science—that are even more complex than historical precedents because they enable not just far more powerful capabilities to design domains external to humans but also the potential to design individual humans themselves. Understanding the implications of this sociotechnical landscape for industrial ecology suggests profound theoretical challenges as well as important new areas of research.